The goal of this project is to develop a novel peptide therapeutic, GLP-1-5, for the treatment of obesity. Today, one-third of adults in the United States (US) are obese, with an estimated annual healthcare burden of approximately $150 billion. By 2030, it is projected that over 50% of adults in the US will be obese with a yearly healthcare burden expected to exceed $860 billion, which will account for approximately 18% of the total US health-care costs. GLP-1 is a glucoincretin hormone that augments glucose-dependent insulin secretion. We discovered that the administration of GLP-1(9-36)amide, the major form of GLP-1 present in the circulation, leads to the formation of two peptides; a nonapeptide, GLP-1(28-36)amide (referred to as GLP-1-9) and a pentapeptide, GLP-1(32- 36)amide (referred to as GLP-1-5) in mouse plasma within 5 minutes. Both of these peptides are the product of GLP-1 cleavage by neprilysin, and both appear to act by a different mechanism than existing obesity drugs, including that of GLP-1 receptor agonists. The peptides appear to enter cells and modify mitochondria function to increase basal energy expenditure via uncoupling of respiration (oxidative phosphorylation). Recently, we demonstrated in mice fed a high-fat diet that infusions of GLP-1-9 increase BEE (Tomas and Habener, manuscript in preparation) and inhibits weight gain. We have now demonstrated that infusions of GLP-1-5 also curtail weight gain and diminish fat mass in mice fed a high-fat diet through increased BEE. While both peptides appear to act through the same novel mechanism, GLP-1-9 is poorly soluble in physiological solutions and considered unsuitable for formulation, making GLP-1-5 a better candidate for clinical development. Our goal is to position GLP-1-5 as a novel therapeutic that functions physiologically as a weight reducer for the treatment of obesity. To our knowledge, no approved drug on the market or in clinical development is able to induce energy expenditure and is the product of a natural protein, in this case GLP-1. The proposed Phase 1 has three key objectives: 1) determine the optimal dose of GLP-1-5, in comparison to the GLP-1 receptor agonist GLP-1(7-36) amide, for maximum BEE and weight loss in obese mice; 2) validate that body temperature and muscle strength are maintained in obese mice with increased GLP-1-5 dosing; 3) establish a reliable potency bioassay to validate the integrity of GLP-1-5 bioactivity.